A goal of our research is to characterize genetic organization in the domestic cat and to develop genomic resources facilitating and establishing Felis catus as a useful animal model contributing to our understanding of human hereditary disease analogues, neoplasia, genetic factors associated with host response to infectious disease and mammalian genome evolution. In order to map and characterize genes associated with inherited pathologies in the domestic cat which mirror inherited human conditions we have focused on increasing microsatellite density in the genetic map of the cat to increase resolution for mapping of genes associated with inherited and infectious disease. The following projects are directed toward these goals: (1) Increasing marker density in the genetic recombination map of the cat: An additional 2000 dinucleotide repeat loci have been isolated, sequenced and characterized from a (dCA.dGT) enriched library of domestic cat genomic DNA. Primers have been designed for amplification of 800 loci; 500 loci have been tested for product performance and locus heterozygosity in a panel of outbred domestic cats. Generation of 3 cM genetic linkage map in the cat which will be applied immediately toward mapping in disease pedigrees: spinal muscular atrophy pedigree, progressive retinal atrophy. (2) Mapping of additional microsatellite loci in an RH map of the cat: Four hundred microsatellite loci have been genotyped in a cat rodent radiation hybrid panel and physically mapped (2 point linkage) relative to 834 previously mapped markers, including 600 Type I coding genes and 234 microsatellite markers. The new markers are well distributed across the cat's 18 autosomes and X and Y chromosome. (3) A generation of a "CEPH" family of cats has been created with permanent cell lines for ongoing genetic analysis: Permanent cell lines have been established from cat pedigrees at the NIH animal facility segregating for coat color variants. The 98 kittens generated in a coat color mapping project provide 198 informative meioses, used in generation of a high resolution genetic linkage map of the cat and provide permanent cell lines for fine scale mapping within genomic regions of interest. (4) On-going construction of a domestic cat genetic linkage map with increased mapping resolution: Six hundred microsatellites are currently being genotyped in the multigeneration 260 member Nestle-Purina domestic cat pedigree. (5) Mapping and characterization of genes associated with inherited disease pathology in cat pedigrees: Degeneration of lower motor neurons in the spinal cord that causes neurogenic muscle atrophy, the so-called spinal muscular atrophies (SMA), are a large category of inherited disorders, but the molecular basis or linkage of a genetic locus has been established in only a few. The SMAs are categorized generally by pattern of inheritance and by distribution of muscle atrophy. In collaboration with John Fyfe (Michigan State), who maintains a pedigree of Maine Coon cats segregating for SMA Type III, we are investigating the molecular basis of this new and unique model of inherited motor neuron degeneration in cats. Linkage analysis of polymorphic microsatellites linked to candidate genes has indicated that disease phenoytpe is unlinked to the feline SMN locus. A full genome scan of microsatellites is being performed in his affected pedigree. Once linkage is established to a cat chromosome, comparative gene mapping will be utilized to identify candidate genes for the pathology. (6) Additionally, development of an STR forensic typing system for genetic individualization of domestic cat specimens: In 1997 we contributed to the establishment of a legal precedent for employing genetic individualization of animal tissue in homicide cases utilizing 10 microsatellite markers. Under support from the National Institutes of Justice, we are continuing this research in developing a microsatellite forensic typing system and genetic database of cat breeds for genetic individualization of cat specimens. A forensic panel of 11 tetranucleotide repeat loci has been isolated from a tetranucleotide enriched library, mapped in the RH map, examined for Mendelian inheritance and characterized in a sample collection of 230 cats representing 29 cat breeds. A robust multiplex amplification protocol utilizing 0.7 ng of DNA has been developed, validated and is currently being genotyped in a panel of 1300 cats representing 37 recognized breeds in the United States. Sample collection of the 1300 cats was also an integral part of the project.